Foxp3+ regulatory T cells, or “Tregs” are fundamental in controlling various immune responses in that Tregs can rapidly suppress the activity of other immune cells. In particular, Tregs are crucial for maintaining tolerance by downregulating undesired immune responses to self and non-self antigens. For instance, Treg defects have been discovered in patients with multiple sclerosis (MS), type I diabetes (T1D), psoriasis, myasthenia gravis (MG) and other autoimmune diseases. Similar links may also exist for atopy and allergic diseases. For all these diseases reports exist pointing to a reduced in vitro immune suppression of the patient's Treg cells. This has led to an increasing interest in the possibility of using Tregs in immunotherapy to treat or prevent, autoimmune diseases, allergies and transplantation-related complications, such as graft rejection or graft-versus-host disease (GvHD) (1).
Methods for the isolation of human CD4+ Foxp3+ Treg cells are known. All of the hitherto described methods for isolation of human Foxp3+ Treg cells employ positive selection of Foxp3+ Treg cells based on cell surface markers of Tregs. That is, the Foxp3+ Treg cells are isolated by using antibodies for Treg associated cell surface markers, mostly CD25+ and CD127−. Yet most cell surface markers of Tregs, such as CD4 and CD25, are not restricted to Tregs. For instance, the commonly employed CD25 is not present on all Foxp3+ Treg cells and is also expressed by effector and memory CD4+ T cells. Consequently, disadvantage of current methods is the contamination of the isolated Treg subsets with effector T cells. When employing markers such as CD25 these contaminations can be significant as up to half of the isolated CD4+ cell population can be comprised of effector T cells. Moreover, it should be further highlighted that there is no method known to isolate CD8+ Foxp3+ Tregs since research has focused on the CD4+ Foxp3+ Tregs. Thus, specific cell surface marker has not yet been identified. However, CD8+ Foxp3+ Tregs also represents an efficient approach for maintaining or inducing tolerance (2).
Hence, there is a particular need for methods and kits/compositions useful for isolating Foxp3+ Treg cells with high degree of purity, and simultaneously CD4+ and CD8+ Foxp3+ Tregs, which are virtually free from CD4+ and CD8+ effector T cells in order to obtain an isolated and purified population of Foxp3+ Treg cells which may be then expanded and pulsed with antigens of interest in order to efficiently induce antigen-specific immune tolerance. Such primed population of Foxp3+ Treg cells is particularly of interest in the fields of autoimmunity, allergy, transplantation, treatment with therapeutic protein and gene therapy, to avoid degradation of self or therapeutic molecules/tissues by the immune system and in cancer to determine the prognosis of a subject suffering from a cancer.